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Regulation of CK2 by phosphorylation and O-GlcNAcylation revealed by semisynthesis

Nature Chemical Biology volume 8, pages 262–269 (2012)Cite this article

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Abstract

Protein serine-threonine kinase casein kinase II (CK2) is involved in a myriad of cellular processes including cell growth and proliferation through its phosphorylation of hundreds of substrates, yet how CK2 function is regulated is poorly understood. Here we report that the CK2 catalytic subunit CK2α is modified by O-linked β-N-acetyl-glucosamine (O-GlcNAc) on Ser347, proximal to a cyclin-dependent kinase phosphorylation site (Thr344). We use protein semisynthesis to show that phosphorylation of Thr344 increases the cellular stability of CK2α by strengthening its interaction with Pin1, whereas glycosylation of Ser347 seems to be antagonistic to Thr344 phosphorylation and permissive to proteasomal degradation. By performing kinase assays with site-specifically phospho- and glyco-modified CK2α in combination with CK2β and Pin1 binding partners on human protein microarrays, we show that the kinase substrate selectivity of CK2 is modulated by these specific post-translational modifications. This study suggests how a promiscuous protein kinase can be regulated at multiple levels to achieve particular biological outputs.

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Figure 1: Preparation of semisynthetic CK2α proteins.
Figure 2: Cellular stability of CK2α.
Figure 3: CK2 interactions with Pin1.
Figure 4: Role of O-GlcNAc modification in CK2α.
Figure 5: Substrate selectivity for CK2.

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Acknowledgements

We thank D. Schwarzer, L. Szewczuk, S. Taverna and Y. Zhang as well as the Johns Hopkins University School of Medicine Microscope Facility for advice and assistance and the US National Institutes of Health (CA42486, GM62437, RR020839) for support.

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Authors and Affiliations

  1. Department of Pharmacology and Molecular Sciences, The Johns Hopkins University School of Medicine, Baltimore, Maryland, USA

    Mary Katherine Tarrant, Hee-Sool Rho, Yu Lin Jiang, Christopher Gross, Jeffrey C Culhane, Gai Yan, Yoshitaka Ichikawa, Jun Seop Jeong, Heng Zhu & Philip A Cole

  2. The Center for High-Throughput Biology, The Johns Hopkins University School of Medicine, Baltimore, Maryland, USA

    Hee-Sool Rho, Jun Seop Jeong, Seth Blackshaw & Heng Zhu

  3. Department of Ophthalmology, The Johns Hopkins University School of Medicine, Baltimore, Maryland, USA

    Zhi Xie & Jiang Qian

  4. The Center for Human Immunology, US National Institutes of Health, Bethesda, Maryland, USA

    Zhi Xie

  5. Department of Chemistry, East Tennessee State University, Johnson City, Tennessee, USA

    Yu Lin Jiang

  6. Department of Biological Chemistry, The Johns Hopkins University School of Medicine, Baltimore, Maryland, USA

    Tatsuji Matsuoka, Natasha Zachara & Gerald W Hart

  7. Biologics Research Laboratories, Kasai R&D Center, Daiichi-Sankyo Co. Ltd., Tokyo, Japan

    Tatsuji Matsuoka

  8. Department of Chemistry, Virginia Tech, Blacksburg, Virginia, USA

    Felicia A Etzkorn

  9. Department of Neuroscience, The Johns Hopkins University School of Medicine, Baltimore, Maryland, USA

    Seth Blackshaw

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Contributions

P.A.C., M.K.T., G.W.H., H.Z., Y.I., C.G., Y.L.J. and J.Q. conceived of the research and planned the experiments. M.K.T., H.-S.R., Z. X., C.G., Y.L.J., N.Z., G.Y. and T.M. performed the experiments. All authors contributed to data analysis and interpretation. J.C.C., F.A.E., J.S.J. and S.B. prepared key reagents. M.K.T. and P.A.C. wrote the manuscript with the support of all of the authors.

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Correspondence to Philip A Cole.

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G.W.H. receives royalties for the sale of GlcNAc-specific antibody.

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Supplementary Methods and Supplementary Results (PDF 4496 kb)

Supplementary Data Set 1

Excel file with the relative signal intensities for individual protein substrates for each of the CK2 enzyme conditions are shown. (XLS 8263 kb)

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Tarrant, M., Rho, HS., Xie, Z. et al. Regulation of CK2 by phosphorylation and O-GlcNAcylation revealed by semisynthesis. Nat Chem Biol 8, 262–269 (2012). https://doi.org/10.1038/nchembio.771

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